The foregoing examples of the related art and limitations related therewith are intended to be illustrative and not exclusive. Other limitations of the related art will become apparent to those of skill in the art upon a reading of the specification and a study of the figures.
Liver disease has become one of the most common chronic illnesses, affecting tens of millions of people in the developed world, resulting in lifetime suffering and huge costs to the medical system. Viral hepatitis C(HCV) is one of the leading known causes of liver disease in the United States. It is a common cause of cirrhosis and hepatocellular carcinoma (HCC), as well as the most common reason for liver transplantation. At least 4 million people in the United States are believed to have been infected with HCV, making HCV the most common chronic blood-borne infection nationally. Treatment of HCV has been successful in up to 60% of the cases, depending upon factors such as genotype, ethnicity, co-infections and other risk factors. In addition to the well-known chronic liver diseases due to viral hepatitis C (HCV), there is an increasing population with chronic liver diseases due to alcohol, autoimmune diseases, obesity and diabetes type 2 (associated with metabolic syndrome). Nonalcoholic fatty liver disease (NAFLD) has become a common chronic liver condition due to obesity and diabetes mellitus, affecting almost a quarter of the general population in the United States. The trend of a rise in obesity in the western world is increasing annually. NAFLD includes a spectrum of liver conditions, ranging from simple steatosis (fat accumulation), also referred to as nonalcoholic fatty liver (NAFL), to non-alcoholic steatohepatitis (NASH) disease, which is associated with liver injury. NASH may progress from fibrosis to cirrhosis, as a consequence of the distortion of the normal liver architecture that interferes with blood flow through the liver. Cirrhosis can also lead to an inability of the liver to perform its biochemical functions, resulting in complications that cause liver failure and liver cancer. Then, transplantation would be the only feasible solution, and in many cases, even transplantation is not an option. Currently, there are several treatments in the pipeline for NASH, but there is no known approved and effective treatment currently available.
Breath tests that are based on monitoring the 13CO2, which is a by-product of metabolization by the liver of 13C labeled substrates, have been proposed as a tool for evaluation of liver function. Previously available tests for liver diseases generally involve drastically invasive procedures, and are therefore much less patient compliant than simple breath tests. Such procedures include biopsies of organs suspected of malfunction, blood tests and imaging technologies. It may take many years, if at all, until liver biopsy will be fully replaced. Although a biopsy is considered to yield reliable results, it is not the optimal tool for patient management since it is highly invasive, expensive, requires day or overnight hospitalization of the patient and is very sensitive to sampling and analysis errors.
Blood tests for the detection of antibodies to suspected bacteria/virus and blood biochemistry tests include standard serum tests and tests following ingestion of suitable compounds. In any event, the blood tests do not specifically diagnose and distinguish NAFLD and NASH from other liver diseases. Most notably, the new serum tests (such as but not limited to FibroTest™) aim at correlation to fibrosis but have difficulties in detecting small changes in liver condition, which are needed for a genuine follow up. None of them have been adapted for use in routine clinical practice yet. They also suffer from the disadvantages of being performed at a central lab, thereby eliminating the economic benefit from the clinic.
Current imaging technologies, including ultrasound, Computed Tomography (CT), X-ray and Magnetic Resonance Imaging (MRI) cannot distinguish NAFLD from NASH. The new Fibroscan (ultrasound) test is not as effective with obese patients (a very significant (and growing) sector of the population), nor does it provide data on inflammation in the liver, nor does it provide information on actual liver function. Most imaging solutions (besides simple ultrasound which is useless for detecting fibrosis) do not exist at the standard medical clinic, and thus such solutions necessitate that the patients leave the physician's clinic, which has its drawbacks, such as location, long waiting period, lack of economic benefits, and more. A basic test in any of these devices (CT, X-ray and MRI) is expensive to highly expensive. Furthermore, there are other disadvantages to the previously used tests, such as the fact that they rarely give real time information about the organ function or status being observed. In some cases, such as in the case of blood tests for antibodies of bacterial infections, they give historic results which may have no current therapeutic relevance, since antibodies to a particular bacterium can remain in the body for up to 2 years from the date that the infection has been eradicated.
Moreover, the liver is an organ that has a very high metabolic capacity reserve. It is well known that a small part of a standard liver mass is sufficient to accomplish its physiological tasks. This poses a challenge when the liver has to be evaluated. Ideally the physician would like to get a quantitative evaluation of the liver mass, percentage of the cells that are functioning normally, or any other related parameter.
Furthermore, it is well known that the liver performs many tasks, and thereby it is difficult to assess all of its functions with a single test. Furthermore, there are many factors that result in high intra- and inter-patient variability. Finally, different disease etiologies may impact different functions of the liver.
The use of two breath tests has been proposed to provide a more accurate picture of the liver diseases. It has been demonstrated that accuracy of evaluation can be improved by using more than one substrate.
Methacetin, also known as N-(4-Methoxyphenyl)acetamide, p-Acetanisidine, p-Acetanisidine and [N-(4-methoxyphenyl)ethanamide] is a compound having the formula:

Methacetin may be utilized for the evaluation of liver functional capacity and/or the extent of liver injury. The biochemical basis for the evaluation of functional capacity is that the compound is metabolized by a cytochrome p450 enzyme expressed in normal liver cells (hepatocytes). Diseases of the liver that cause a loss in functional mass and/or impact the metabolic function of hepatocytes are associated with and may be correlated to a loss of capacity to metabolize methacetin.
One of the most common methods for determining the rate of metabolism of methacetin is to analyze the rate of metabolism of the methoxy group (CH3O) of methacetin to carbon dioxide, which is excreted in exhaled breath. To distinguish the carbon dioxide derived from methacetin from all other sources of carbon dioxide, the methoxy group is labeled with 13C, a stable isotope of carbon. Thus, all the CO2 derived from methacetin will contain 13C(13CO2) in contrast to all other sources of CO2, which will contain approximately 99% 12C, and 1% 13C, the naturally abundant isotope. Thus, the rate of excretion of 13CO2 (normalized to 12CO2) above background following the administration of methacetin-methoxy-13C indicates its rate of metabolism, which relates to the hepatic cell “health” and to the functional mass of the liver.
In common tests 75 mg of methacetin-methoxy13C is dissolved in 50-200 ml of water and taken orally, following which the excretion rate of 13CO2 in exhaled air (breath testing {BT}) is determined at intervals of 15 minutes up to 2 hr. It has been reported that individuals with well-established cirrhosis have a statistically significant reduction in the rate of metabolism of methacetin, but considerable overlap exists between a group of volunteers having normal liver function and those with milder stages of potentially progressive liver disease and/or the degree of liver injury.
The probable causes for the wide variation in the methacetin breath testing within the normal population, which makes it difficult to distinguish it from the population with mild loss of functional capacity, need to be addressed and overcome as well as the intra-patient test variability. There is a need for a modified test that would enhance the usefulness of the methacetin in breath test for evaluating liver functional capacity and/or hepatic injury or health.
Octanoic acid, a medium-chain fatty acid and salts thereof undergo a metabolic process in the mitochondria of the liver cells. These compounds may be used in the assessment of hepatic mitochondrial β-oxidation. There is a need in the art for a test that would allow accurate evaluation of hepatic related conditions.